First Report of Macruropyxis fraxini Causing Rust Disease on Microstegium vimineum in China

Stiltgrass (Microstegium vimineum (Trin.) A. Camus) is an annual Poaceae weed with a broad native range throughout East Asia. Stiltgrass is an invasive grass that is distributed in more than 15 provinces in China, posing a major threat to native biodiversity and restoration efforts in introduced areas. Stiltgrass often forms dense near-monocultures in forest understories and riparian areas where it disrupts forest succession, nitrogen cycling, and alters native communities (Stricker et al. 2016). In August 2018, M. vimineum with rust disease were observed near the roadside (26.759482 °E, 114.283519 °N) in Jinggangshan City, Jiangxi Province, China. Diseased plants were observed at a 2 × 10 m shady location with lesions on leaves and stems, disease incidence was over 90% (n=100). Sixty disease samples were collected to confirm the pathogen. Early symptoms on the upper leaf surfaces consisted of rust pustules, which were circular, subcircular to irregular, orange to dark-orange, crust-like, and granular. At later stages, lesions coalesced, spreading all over the plant, causing severe defoliation. Uredinia were predominantly formed on the upper surface of leaves and young stems but rarely also found on the abaxial leaf surface, exposed, yellow to yellow-orange, and 0.2-0.5 mm in diameter, occasionally reaching 0.9-1 mm, surrounded by purple lesions (n=30) (Olympus SZX7). Telia were predominantly formed on the lower surface of leaves, stems, exposed, chestnut-brown to dark-brown. Urediospores were nearly spherical, oval or obovate, light yellow, 18-23 μm × 20-26 μm, cell wall is about 2-2.5 μm (n=200) (ZEISS AXIO Imager. M2). Teliospores were ellipsoid, 37-55 μm × 25-36 μm, 2-celled, inner wall brown, 4-5.5 µm thick, outer wall hyaline, smooth, germ pores 2-4 per cell; pedicels were hyaline, composed of cell walls with loss of cytoplasm, 4.5-6.5 µm wide, and up to 160 µm long (n=200) (ZEISS AXIO Imager. M2). Pycniospores, aeciospores and basidiospores were not observed in this study. The telial morphology features were consistent with those reported of M. fraxini, but uredinial stages were not observed in these studies (Azbukina 1974; Jung et al. 2020). Genomic DNA was extracted from a representative specimen (JGS-1) and was characterized by PCR amplification and sequencing of 28S rDNA using the primer pair NS1 and NS4 (Aime 2006). The 1094-bp sequence (Genbank: ON739170) shared 99.18% nucleotide identity with M. fraxini (Genbank: KP858144). The internal transcribed spacer (ITS) region was sequenced by rust fungal primer pairs ITS4rust and ITS5u (Pfunder et al. 2001). The 564-bp sequence (Genbank: ON739169) shared 99.12% nucleotide identity with M. fraxini (Genbank: KP858145), which was consistent with the morphological features observed. To complete Koch's postulates, plants were inoculated by brushing a urediniospore suspension (1 ×105 spores/ml) onto the leaves, placed in a plant growth chamber (25℃, 8 h/d of dark, 30℃, 16 h/d of light, 8000 lux of light intensity, RH ≥ 90%). Urediniospores were formed on the leaf surface 7 to 10 days after inoculation, and all infected plants showed symptoms similar to those observed in the field, along with spores, whereas the control plants remained symptomless. Host range tests showed that rice, wheat, barley, sorghum, maize, cotton, peanut and rape were resistant to M. fraxini but soybean and peas were susceptible. More research is needed to determine whether this pathogen can be a biocontrol agent for stiltgrass, such as exploring the potential impact of this rust pathogen, expanding host range tests, and finding its alternate hosts. To the best of our knowledge, this is the first report of rust disease on stiltgrass caused by M. fraxini in China.